Our long-term goal is to discover novel therapeutic targets for pelvic pain. Pelvic pain, bladder pain specifically, is also the cardinal symptom of interstitial cystitis/bladder pain syndrome (IC/BPS) where most patients show no underlying bladder infection, pathology, or injury. The local and central mechanisms that mediate pelvic pain are not fully understood. There is an urgent need to correct this knowledge gap so as to develop effective novel therapeutic targets for chronic pelvic pain. Our overall objective in this application is to determine the mechanisms of MIF- mediated bladder hyperalgesia. Our central hypothesis is that MIF mediates bladder hyperalgesia through local (bladder) and central (spinal) mechanisms. We plan to test our central hypothesis by pursuing the following three specific aims: 1. Identify urothelial MIF receptors that mediate bladder hyperalgesia. Based on the published data referred to above, our working hypothesis is that released MIF acts at urothelial MIF receptors to elicit urothelial HMGB1 release and bladder hyperalgesia. 2. Identify downstream targets of MIF/HMGB1 that mediate bladder hyperalgesia. Our working hypothesis, based on our published and preliminary evidence, is that blocking downstream signaling pathways activated by MIF/HMGB1 will prevent bladder hyperalgesia. 3. Identify spinal mechanisms that mediate persistent bladder hyperalgesia. Based on our preliminary data, our working hypothesis is that increased production and release of spinal MIF activates increased release of spinal HMGB1 to maintain bladder pain. In order to accomplish these aims we will conduct studies using rodent models of bladder pain and in vitro studies using human urothelial cells. We will examine organ-level and spinal-level mechanisms of bladder pain while focusing on treating persistent bladder pain. We will also identify novel lumbosacral peptides/proteins associated with persistent bladder pain. Succesful completion of these studies is expected to have a positive impact because it is probable that the identified components of the mechanism will provide new peripheral and/or central targets for therapeutic interventions in conditions of bladder pain.